Embossed film bioprocessing containers and integrity testing of bioprocessing containers

ABSTRACT

A testing method that includes the steps of evacuating air from a container to a negative atmospheric pressure, the container being a collapsible, flexible container, and comprising at least two opposing flexible walls, wherein a surface of at least one of the walls internal to the container comprises a plurality of channels or recessed features on said at least one wall and monitoring a mass flow or a state of vacuum so as to determine the integrity of the container. The container can be of any size or conformation, with or without attached fittings.

RELATED APPLICATIONS

The present application claims the benefit of priority of U.S.Provisional Patent Application No. 62/567,266, filing date Oct. 3, 2017,the entire content of which is incorporated herein in its entirety.

BACKGROUND Field of the Technology

Provided herein are materials useful for manufacturing bioprocessingcontainer(s) and methods of integrity testing bioprocessing containers.Film materials having textured or embossed patterns, for manufacturingdisposable, single-use containers and systems useful for containing,transporting, mixing and/or processing biological liquids and/orsolutions, are disclosed. More particularly, integrity testing of suchcontainers, including small and large volume containers of varyingdimensions and complexity, are disclosed.

Description of the Related Art

Fluids used in industries, such as bioprocessing and pharmaceuticalfields, have been traditionally processed in systems that use stainlesssteel containers. After each use, these stainless-steel containers aredisassembled, cleaned, sterilized and reassembled before reuse toprovide sterile components. The cleaning and sterilization proceduresare time-consuming, expensive and cumbersome. If the containers are notproperly cleaned, reassembled, and sterilized, the containers can becontaminated and compromise the fluid contents in a subsequent use.Furthermore, the cleaned and sterilized containers typically need to bevalidated before use, adding expense and time burden to the process.

To provide greater flexibility in processing and to reduce time spent onvalidating equipment, manufacturers have developed disposablepre-sterilized or on-site sterilizable containers that are used once anddisposed. Flexible or collapsible bags are an example of suchsingle-use, disposable containers. Various attachments on the single-usecontainers, such as tubing, port fittings, connectors, mixing elementsand the like, may also be disposed with the container.

Maintaining sterility of the single-use container or bag is of vitalimportance, particularly for the food, beverage, and pharmaceuticalindustries. Contamination poses serious health and environmental risks.Furthermore, loss of product through leaks can cause significanteconomic loss.

Stainless steel containers are robust and are not readily punctured ortorn. In contrast, flexible containers, which can be made from plasticor silicone materials, are more susceptible to being torn or punctured.Therefore, the integrity of flexible containers is tested to check fordefects that would compromise the system.

Major integrity failures in single-use bioprocessing containers areoften readily apparent and can be identified visually. Integrityfailures include seam failures, rips, and/or visible puncture holeswithin the container or film(s) of which the container is manufactured.However, smaller defects, such as microscopic holes or tears, cannot bedetected by mere visual inspection. These smaller defects represent agreater risk since, in addition to leaking, they can permit the passageof microorganisms, particles, fluids or other unwanted materials intothe system, which may go undetected for a period of time during use.Discovering contamination after a bioprocessing run has started mandatesthat the batch be discarded, and the process started again, which wastestime and resources.

Current methods of non-destructive integrity testing inflate thebioprocessing containers, such as bags, with a gas and test for leak ofthat gas. Leaks can be measured in different ways. For example, in thepressure decay integrity test, the bag is inflated with air to a setpressure, stabilized for a predetermined time to counteract stretchingand adiabatic effects of filling, and monitored for decay in thepressure in the bag. Loss of pressure indicates a leak.

The pressure decay integrity test is relatively reliable for smallcontainers, but not for larger containers. Increasing pressure in alarger flexible container during a pressure decay test subjects thecontainer to much greater strain on the material than for smallercontainers and is more susceptible to burst. This is because the hoopstress in the wall of the container increases with the radius.Therefore, a larger container that has the same wall thickness as asmaller bag is subject to greater stress under the same pressure.

For example, a 2-liter flexible film single-use process container (forexample, MOBIUS® PureFlex™ bags, manufactured by MilliporeSigma,Burlington, Mass., USA) can be safely inflated to 1.5 pounds per squareinch (PSI) (10.3 kPa). At this pressure, a hoop stress of roughly 300PSI (2068 kPa) will be developed in the wall of the bag. This isacceptably below the 1360 PSI (9377 kPa) yield stress of the film and,therefore, will not damage the bag. At these conditions, with asensitive pressure transducer, a 30 μm defect can be found within 5minutes. However, sensitivity is lost significantly with increasing sizeof the container. For example, if the container or bag has a 20-litervolume, a test pressure of 1.5 PSI (34.5 kPa) develops a hoop stress of1500 PSI (10342 kPa), which typically exceeds the yield stress of thefilm. For a 20-liter sized bag, the test pressure would have to bereduced to 0.6 PSI (4.1 kPa) to keep the hoop stress well below theyield stress. At this lower test pressure and higher bag volume, onlylarger defects of at least 170 μm are expected to be identified in 5minutes. In other words, as the container or bag volume increases, thetest pressure needs to decrease and, therefore, the test sensitivity,unfavorably, decreases commensurately.

Another problem with using pressure decay methods to test bags is thatthe flexible nature of disposable containers permits expansion of thefilm material itself, which in turn appears as pressure decay, leadingto a false reading of an integrity failure. The presence of an actualdefect in a large, flexible container can also be missed during apressure decay test because the difference in pressure decay caused bythe leak is masked by the pressure decay caused by the stretching of thefilm material, and/or the seams of the film materials, itself.

Past attempts to minimize the film material stretch of large flexiblecontainers in the pressure decay test, thereby reducing the hoop stresseffect, include methods wherein the integrity test is conducted with thecontainer restrained between two rigid plates. While this reduces theamount of material stretch during the pressure decay test, this testsuffers from masking defects located adjacent to the rigid plates, i.e.,the plates seal the defects. Consequently, the problem remains that adefective container may erroneously pass the integrity test.

To avoid the problem of restraining plates masking actual defects, anadditional porous material needs to be placed adjacent to the plates.The porous material acts to prevent defects from being sealed offagainst the plate. This is, however, unsuitable for containers that arenot sufficiently flat. For example, the pressure decay test of largebioprocessing containers with attached fittings, such as tubing,connectors, etc., is not suitable for use with constraining plates. Theattached fittings compromise the ability of the rigid plates to lieflush with the container, thereby defeating the purpose of the plates,e.g., to minimize material stretch during pressurization. Additionally,disproportionate pressure may also be exerted at points of contactbetween the constraining plates and fittings on the container, whichmay, in turn, cause the rupture of a join or seal between the fittingand the container.

Other integrity testing methods have included the use of Helium gas. Inthis test, the container being tested is connected to a Helium sourceand placed into a sealed, rigid vessel with an outlet. The air is pulledfrom the rigid vessel through the outlet, and Helium gas is injectedinto the container being tested. If the container has a defect, Heliumescapes from the container into the rigid vessel and can be detectedusing mass spectroscopy. However, volumetric leak flow rate depends ongas viscosity, and Helium has a higher gas viscosity than a gas such asNitrogen. Consequently, a container with a defect will leak less withHelium than Nitrogen under the same differential pressure. As such,leaks can take longer to detect or require higher pressures to detect.Furthermore, Helium can diffuse through many materials, includingsilicone which is often used in flexible bioprocessing containers.Consequently, detecting Helium in the test may be the result of adefect, or, alternatively, a false positive from Helium diffusionthrough the film material itself.

In view of the foregoing, an integrity test that overcomes the abovedeficiencies would represent an advance in the art.

SUMMARY

A testing method that includes the steps of evacuating air from acontainer to a negative atmospheric pressure, the container being acollapsible, flexible container, and comprising at least two opposingflexible walls, wherein a surface of at least one of the walls internalto the container comprises a plurality of channels on said at least onewall and monitoring mass flow so as to determine the integrity of thecontainer. The container can be of any size or conformation, with orwithout attached fittings.

Integrity tests according to embodiments of the present disclosure arehighly sensitive, sensitive enough to detect microscopic defects in anysize or conformation of vessel, whether a single-use disposabletwo-dimensional (2-D) or three-dimensional (3-D) bag, with or withoutattached fittings. Some embodiments disclosed herein comprise methodsfor determining the integrity of a collapsible, flexible bioprocessingcontainer that eliminates variation in testing due to stretching ofmaterials and films during testing. Some embodiments disclosed hereincomprise methods for determining the integrity of a collapsible,flexible bioprocessing container that eliminates the creation of defectsdue to stretching of materials and films during testing

A highly sensitive and rapid method of determining the integrity of acollapsible, flexible bioprocessing container is disclosed. The methodincludes evacuating air from the container to a negative atmosphericpressure, the container being a collapsible, flexible bioprocessingcontainer, and comprising at least two opposing flexible walls, whereinthe surface of the walls internal to the container comprise a pluralityof channels on said walls and; monitoring mass flow so as to determinethe integrity of the container. The bioprocessing container can be ofany size or conformation, with or without attached fittings. In someembodiments, the container comprises at least two opposing flexiblewalls. The surface of at least one wall, one of two opposing walls, orall the walls, that are internal to the container comprise a pluralityof channels. In some embodiments, the plurality of channels is embossedon the wall(s). Alternatively, or additionally, the internal surface ofthe wall(s) is a matte surface. In some embodiments, the collapsible,flexible bioprocessing container is pre-folded. Any embodiment accordingto the disclosure contemplates a container further comprising fittings,tubing, connectors, or any combination thereof.

In some embodiments, an integrity test comprises evacuating air from thecontainer to a negative atmospheric pressure and monitoring a mass flowof air or a vacuum decay, so as to determine the integrity of thecontainer. An increase in mass flow during monitoring indicates thepresence of a leak.

In some embodiments, a method of detecting a 2 micron or larger defectin a collapsible, flexible bioprocessing container is disclosed. Themethod comprises providing a collapsible, flexible bioprocessingcontainer, which container comprises at least two opposing flexiblewalls. The surface of at least one wall, one of two opposing walls, orall the walls, that are internal to the container and further comprisesa plurality of channels. The method comprises evacuating air from thecontainer to a negative atmospheric pressure and monitoring mass flow ofair or vacuum decay. An increase in mass flow and/or a loss in thevacuum indicates the presence of a defect in the container.

In some embodiments, the integrity test comprises evacuating air to anegative atmospheric pressure between approximately −1 (negative) PSI(6.9 kPa) to approximately −14 (negative) PSI (96.5 kPa).

BRIEF DESCRIPTION OF THE DRAWINGS

The drawings are provided to illustrate one or more embodiments of thedisclosure and are not to be construed as limiting the scope of theclaims.

FIG. 1A depicts a textured or embossed pattern having raised featuresand recessed channels on an interior of a flexible film according toembodiments of the disclosure;

FIG. 1B depicts a textured or embossed pattern having recessed featuresand raised channels on an interior of a flexible film according toembodiments of the disclosure;

FIG. 2A depicts a cross section view of the textured or embossed patternof FIG. 1A;

FIG. 2B depicts a cross section view of the textured or embossed patternof FIG. 1B;

FIG. 3 is a graph comparing integrity test results with flexiblecontainers having smooth or embossed walls, with or without an addeddefect;

FIG. 4 is a graph comparing integrity test results of a small (20 L)embossed bag with a 2 μm sized defect, compared to the same bag havingno defect;

FIG. 5 is a graph comparing integrity test results of a larger (200 L)bag with a 2 μm defect, compared to the same bag having no defect;

FIG. 6 is a graph comparing integrity test results of a large (500 L)bag with a 5 μm defect, compared to the same bag having no defect;

FIG. 7 is a graph comparing integrity test results of a 200-liter bagafter use (filled with water and emptied) with a 50 μm defect, comparedto a 200-liter bag with a 50 μm defect before use (not filled withwater), a 200-liter bag having a defect both before and after use; and

FIG. 8 is a schematic diagram outlining an exemplary integrity testingsystem of some embodiments of the disclosure.

DETAILED DESCRIPTION

Traditional pressure-based integrity tests have limitations, especiallyas the volume of the flexible container increases in size (e.g., morethan 20 liters) and in complexity (e.g., 3-D versus 2-D, containerswith, optionally, attached ports, connectors, tubing, and otherfittings). It is to be understood that the terms containers and bags areused interchangeably throughout this disclosure.

Contrary to the traditional integrity tests that need to inflate thecontainer with or without a tracer gas, the present system uses a vacuumto test the integrity of a flexible container. The presence ormeasurement of fluid flow, e.g., air or another gas, following theevacuation of the flexible container indicates the presence of a defect.Similarly, the measurement of a loss of vacuum would also indicate thepresence of a defect, i.e., vacuum decay.

Embodiments of the vacuum integrity test(s) disclosed herein areunexpectedly more sensitive than traditional tests that inflate thecontainers. Furthermore, some embodiments of the vacuum integritytest(s) disclosed herein can detect a defect(s) more rapidly thanpressure decay tests. The vacuum integrity test(s) disclosed hereinavoid stretching of the film material under the inflation pressure,thereby avoiding any risk of rupture or creation of other defect(s).

After traditional inflation-based integrity tests, the container needsto be deflated and prepared for packaging and shipping. Unfortunately,this handling process can inadvertently introduce defects in theflexible container through improper handling, abrasion, creasing, and/orfolding. Conveniently, the vacuum integrity test according toembodiments of the disclosure can be performed on bioprocessingcontainers and systems that are already folded and ready for finalpackaging and shipping. This method(s), therefore, avoids introducingthe defects caused during handling of the container after traditionaltests. Also, final packaging and shipping can be performed following,e.g., immediately, successful testing, saving time, logisticalconsiderations, expenses, and like operational concerns.

The vacuum integrity test(s) disclosed herein is also faster thantraditional inflation-based integrity tests, which facilitates fasterquality control and higher output for manufacturers and fastervalidation by end-users.

Embodiments of the vacuum integrity test(s) disclosed herein uses avacuum to evacuate air from the interior of a flexible container havingthe specific properties described herein. Once the container issufficiently evacuated, further air flow can only be the result of adefect or leak in the bag. Similarly, once the container is sufficientlyevacuated, the presence of a loss of vacuum, i.e., vacuum decay, canonly be the result of a defect or leak in the bag. Using a vacuum toremove air from traditional containers having flat, non-textured walls,or non-embossed walls is not efficacious. In this situation, when vacuumis applied to a traditional container, the interior faces of thecontainer are brought into intimate contact with each other (i.e., whenthe container collapses). Pockets of air cannot be fully evacuated dueto the sealing effect of the collapsed walls, occurring for either 2Dand/or 3D flexible containers. In this state, defects are masked,because of the intimate contact of the interior walls. A hole/defect inone wall, will be sealed against the other wall, thereby blocking anyair flow through the hole/defect. Also, any defect in an air pocketbehind the sealed surfaces are similarly masked. The presence of thehole/defect is, therefore, undetectable.

Some embodiments of the method disclosed herein avoid these problems viathe use of a textured or embossed film in construction of the flexiblecontainer. Specifically, embodiments of the disclosure comprise bags orbiocontainers wherein the interior walls of a container or bag areproduced with an embossed or textured pattern. The high points, e.g.,raised areas, of any pattern, such as raised squares having loweredchannels therebetween, according to embodiments of the disclosure, actas spacers that maintain a small separation between the interior wallsof the container when it is in its collapsed state. The space, e.g.,channels thereby produced between the raised areas, prevent defects frombeing sealed off by the opposing interior wall of the container duringvacuum testing. This small, open channel(s) allows air flow coming froma defect to occur under vacuum testing. This air flow is detected by asensitive mass flow meter that is in-line between the flexible containerbeing tested and a reference vacuum tank. In some embodiments, squareareas are recessed into a surface of the textured or embossed film,wherein the raised areas are rails.

By way of example, and not limitation, FIG. 3 shows the effect of havingan embossed pattern on the interior wall of the container. A containerwith the embossed pattern shows a relatively high air flow when a defectis in place. In contrast, the air flow through a container without anembossed film is significantly lower. Without intending to be limited bytheory, it is believed that the embossed film, which presents as peaksand valleys, prevents the film from sealing against itself, which caninterfere with the identification of defects.

Optionally, the vacuum integrity test can be performed before shipping,after delivery, before use, and/or after use. In this way, the integrityof the disposable, flexible container can be assured when it leaves themanufacturing facility, upon receipt by the customer, before fillingwith ingredient(s) and processing, and after processing is complete.Furthermore, multiple tests can be conducted without the risk ofcreation of defects.

In accordance with certain embodiments, the flexible container isdesigned to receive, hold, mix and/or dispense materials, such asbioprocessing media, buffers, fluids, cells, tissue, and the like.Typically, the flexible container is aseptic or sterile.

The flexible container can be formed from a flexible monolayer film ormultilayer film, the film forming one or more flexible walls. Theflexible film can have a polymeric composition comprising one or morematerials such as polyethylene, including ultrahigh molecular weightpolyethylene, linear low density polyethylene, low density or mediumdensity polyethylene; polypropylenes; ethylene vinyl acetate (EVOH);polyvinyl chloride (PVC); polyvinyl acetate (PVA); ethylene vinylacetate copolymers (EVA copolymers); blends of various thermoplastics;co-extrusions of different thermoplastics; multilayered laminates ofdifferent thermoplastics; or the like. By “different” it is meant toinclude different polymer types such as polyethylene layers with one ormore layers of EVOH as well as the same polymer type but of differentcharacteristics such as molecular weight, linear or branched polymer,fillers and the like. Typically, medical grade and/or animal-freeplastics are used. The film is generally sterilizable such as by steam,ethylene oxide, or radiation such as beta or gamma radiation. Most filmsused in the manufacture of flexible, single-use biological containershave good tensile strength, low gas transfer and are either transparentor at least translucent. Typically, the material is unsupported and/oris weldable. Also, the film used to manufacture the containers is clearor translucent, allowing visual monitoring of the contents of thecontainer. The container is typically provided with one or more inlets,one or more outlets and one or more optional vent passages. One or moreimpeller assemblies can be positioned in the container for mixing thecontainer contents.

In some embodiments, the container may be a disposable, deformable,foldable bag that defines a volume, is sterilizable for single-use, andcapable of accommodating contents, such as biopharmaceutical liquids. Insome embodiments, the container can accommodate one or more mixingdevices partially or completely within the interior of the container.Mixing devices include but are not necessarily limited to impellers,baffles, spargers, and the like, as are known to those in the art. Thevolume can be a closed volume that can be opened, such as by suitablevalves, to introduce a fluid into the volume, and to dispense fluidtherefrom. In some embodiments, the flexible containers can includeports, connectors, tubing, baffles, vortex breakers, additional bags,and other fittings. Containers according to some embodiments disclosedherein are sometimes referred to as bioprocessing systems.

Flexible containers, such as bioprocessing bags, for this vacuumintegrity test(s), disclosed herein, are not limited in size. Indeed,unlike traditional inflation-type integrity tests, this vacuum integritytest is equally suited for very large containers (e.g., at least orabout 2000 liters, 3000 liters, 3500 liters, 5000 liters, or more), aswell as smaller containers (e.g., 50 liters, 10 liters, 5 liters, 1liter, or the like). Similarly, vacuum integrity test(s) describedherein can be applied to flexible containers that are 2-D (e.g.,pillow-shaped bags) or 3-D (cylindrical, cuboid, and/or the like,containers).

The container is bound by one or more walls, typically made of aflexible film. The walls comprise an inside and outside surface relativeto the container. In some embodiments, the container comprises at leastone wall that has an internal surface that is not smooth. In otherwords, the internal surface of at least one wall has a texture. Thetexture provides a plurality of channels on the surface of the wall,which facilitates fluid (e.g., a liquid or a gas) to pass through thechannels. The texture can be any suitable texture such that the textureprovides a plurality of channels. For example, the texture can beirregular, random, patterned, non-patterned, symmetrical,non-symmetrical, repeating or non-repeating. The texture may be matte,embossed, grooved, or otherwise formed on the surface of the film. FIGS.1A-1 B illustrate examples of texture that is embossed on a flexiblefilm; FIGS. 2A-2B illustrate cross section views of the texturesembossed on the flexible films of FIGS. 1A-1 B.

FIG. 1A depicts a textured or embossed pattern having raised features110 and recessed channels 120 on an interior of a flexible film 100according to embodiments of the disclosure. As shown, FIG. 1A depictsraised areas 110 and channels 120, wherein the raised areas 110 areshown as squares. It is to be understood that other geometries arepossible. For example, circular raised areas, rectangular raised areas,rhomboidal raised areas, etc. Also, in some embodiments, the squares,circles, rhomboids, etc., may be recessed from a surface. In suchembodiments, the channels in FIG. 1A would present as raised rails, asdiscussed below. In the example of FIG. 1A, the texture comprises arepeating pattern of raised squares 110, wherein the spaces between theraised squares form a plurality of channels 120, which are optionallyinterconnected, through which a fluid, e.g., a biological fluid beingprocessed or a gas during vacuum integrity testing, can pass. Thetexture can be formed from any shape, not just squares, such as circlesor dots, lines or grooves, a weave pattern, or other such shape. Any ofthe embossing patterns disclosed herein comprise peaks and valleys,i.e., raised areas (e.g., squares) and channels extending between theraised areas. In some exemplary embodiments, the height of the raisedareas is approximately 0.06 mm to approximately 0.013 mm. The width ofthe raised areas may comprise, for example, approximately, 0.05 mm to0.10 mm. It is to be understood that other sizes and dimensions ofembossed areas are within the scope of the present disclosure. It is tobe further understood that the embossed pattern in any film may be thesame as or different from other embossed patterns irrespective of thesize of the bag to be tested using embodiments of the methods disclosedherein. FIG. 2A depicts a cross section view 150 of the textured orembossed pattern of embodiments of FIG. 1A. FIG. 2A more clearlyindicates that the cross section 150 of the flexible film 100 of FIG. 1Acomprises raised features 110, displayed as squares, and recessed areas120, wherein the raised features 110 are disposed on the flexible film100 on an interior of a bag or biocontainer.

FIG. 1B depicts a textured or embossed pattern having recessed features130 and raised channels 140 on an interior of a flexible film 200according to embodiments of the disclosure. The embodiments of FIG. 1Bare substantially the mirror of the embodiments of FIG. 1A. FIG. 2Bdepicts a cross section view 250 of the textured or embossed pattern ofembodiments of FIG. 1B. FIG. 2B more clearly indicates that the crosssection 250 of the flexible film 200 of FIG. 1B comprises raised rails140 and recessed areas 130, displayed as squares, wherein the raisedrails 140 are disposed on the flexible film 200 on an interior of a bagor biocontainer.

In some embodiments, the container comprises at least two opposingflexible walls. The internal surface of each of the two opposing wallsmay each, or individually, comprise a plurality of channels on saidwalls. In at least one embodiment, the internal surface of all walls ofthe container are textured and comprise a plurality of channels. In someembodiments, the texture on opposing walls are non-interlocking witheach other, such that when the opposing walls are adjacent to eachother, the channels are not obscured or blocked by the texture on theopposing wall.

In performing the vacuum integrity test(s), the plurality of channelsformed by the texture allows for the contents of the flexible containerto be evacuated, such as by vacuum suction. In some embodiments, theflexible container undergoes a vacuum until a negative pressure isreached. For example, from approximately −1 (negative) PSI toapproximately −14 (negative) PSI. Once the predetermined negativepressure is attained, the air flow is monitored using a mass flow meter.A high air flow, such as 20% or greater air flow, depending on the airpressure during testing, indicates the presence of a defect. Inexemplary embodiments, 20% greater air flow can be achieved even, forexample, for defects as small as two to five microns in size. Air flowcan be monitored and compared to a suitable control, such as anair-tight non-leaking container, and/or a container with a calibratedsize orifice. Also, one a bag is evacuated, a loss of vacuum can only bethe result of a defect or leak in the bag, i.e., vacuum decay, a stateor condition of a loss of vacuum. In some exemplary embodiments, testingmay occur from a range of approximately −5 (negative) PSI toapproximately −7 (negative) PSI.

Generally, during the test the flexible container is in directcommunication with a rigid steel reference tank. Both the tank andflexible container are evacuated of air. Once the evacuation iscomplete, the system is sealed, and a mass flow meter situated betweenthe reference tank and the flexible senses any air flow from thecontainer to the reference tank.

As described herein, the size of a defect, or a leak, detected comprisesa range of approximately 1-250 microns, and more typically approximately2-10 microns.

In some embodiments, the mass flow is tested for approximately 10minutes or less, in some embodiments, approximately 2 minutes or less,more typically, approximately 20-30 seconds.

FIG. 3 shows a graph comparing the vacuum integrity test on a textured(embossed) film bag with a standard non-textured surface bag. In thisembodiment, a 20-foot long (which may be, for example, 5 inches wide),20-liter bag was integrity tested with artificially introduced defectsof varying sizes. Defects were introduced as either laser-drilled holesof 100 microns and 50 microns (“in film”) or as a 30 micron hole in afitting attached to the bag (“in tube”). An increase in flow rate wasdetected in less than 30 seconds for defects in bags with an embossedfilm, whether on the bag or on the fitting. The relationship between thecontainer and the test sensitivity is indicated in FIGS. 4-6.

FIG. 7 confirms that the vacuum integrity test can be performed post-useof the bag. For example, a container can be filled with fluid (such asfor mixing, bioprocessing, etc.). Following use, confirmation of theintegrity of the container may be desired. Typically, once the containeris drained, some residual fluid may remain. This residual fluid shouldbe prevented from entering the mass flow meter. As such, the residualfluid can be evacuated under vacuum into a water trap before conductingthe vacuum integrity test. By inclusion of valves in the system,contents from the container can be first diverted through the water trapbefore commencing the vacuum integrity test. Filters are furtheroptionally included in the lines.

FIG. 8 is a schematic diagram outlining an exemplary integrity testingsystem 800 of some embodiments of the disclosure. The system may be usedto vacuum integrity test a container, e.g., a biocontainer or a bag,after use, such as after fluid has been handled in the container anddrained. Residual fluid left in the container may be captured in thewater trap before conducting the vacuum integrity test. The water trapsystem is an optional component and may be omitted if no or little fluidis present in the container being tested.

The integrity testing system 800 comprises a vacuum pump 802, a tester804, and a bag 806 to be tested, for example, a bag or biocontainerhaving an embossed film, as described in greater detail above. Thevacuum pump 802, the tester 804, and the bag 806 are connected viaconduits 808. The integrity testing system 800 may further comprise oneor more air filters 810. The integrity testing system 800 may alsofurther comprise one or more water traps 812, according to someembodiments of the disclosure. The integrity testing system 800 mayfurther comprise one or more valves 814, for opening and closing asappropriate. For example, the valves 814b may be closed in conduits 808adjacent to the tester 804. While the valves 814a are open, pulling avacuum permits water to flow and be trapped by a water trap 812.Subsequently, the valves 814b are opened and the valves 814a closed, sothat the bag 806 may be tested in a substantially liquid, e.g.,water-free environment. The integrity system 800 may be used before orafter the bag 806 is used to process, for example, biological liquids.The vacuum pump 802 pulls a vacuum, e.g., from −1 to −14 PSI, from thebag 806, wherein the tester 804 is disposed between the vacuum pump 802and the bag 806. The tester 804 will indicate a flow of air or,alternatively, a loss of vacuum, as described above, if a defect ispresent in the bag 806.

It is to be understood that any of the embodiments of the film(s) tomanufacture a container and/or biocontainer, as described herein, maycomprise a single layer film or a multilayer film. It is to be furtherunderstood that whether a single layer film or a multilayer film, eithermay be textured or embossed. It is further to be understood that theinternal contact layer of any film would be textured or embossed. It isfurther to be understood that any embodiments of the container,biocontainer, or film can be textured or embossed as is known to thosein the art. For example, the single layer film or the multilayer filmcan be embossed during an initial extrusion, after extrusion, e.g., asecondary operation, or can be embossed after all layers, i.e., for amultilayer film, are laminated, calendared, or otherwise adheredtogether.

The integrity test(s) described herein uses a vacuum and consequently isnot limited in the size or shape of the container being tested. Outlets,inlets, ports or other openings in the container or on any attachedfittings, can be sealed or closed prior to the vacuum integrity test.The described integrity test can be universally applied to bioprocessingcontainers and systems and overcomes shortcomings of standardinflation-based integrity tests.

At least some testing methods according to embodiments of the disclosurecomprises evacuating air from a container to a negative atmosphericpressure, the container being a collapsible, flexible container, andcomprising at least two opposing flexible walls, wherein a surface of atleast one of the walls internal to the container comprises a pluralityof channels on said at least one wall and, monitoring mass flow (or theloss of vacuum) so as to determine the integrity of the container, i.e.,the presence or absence of a defect.

At least some testing methods of detecting a 2 micron or larger defectin a collapsible, flexible bioprocessing container, the method compriseproviding a collapsible, flexible bioprocessing container, the containercomprising at least two opposing flexible walls, wherein the surface ofthe walls internal to the container comprise a plurality of channels onsaid walls; evacuating air from the container to a negative atmosphericpressure; and monitoring mass flow, wherein an increase in mass flowindicates the presence of a defect.

Any of the testing methods according to the disclosure may comprisewherein the negative atmospheric pressure is between about −1 PSI to −14PSI. Any of the testing methods according to the disclosure may comprisewherein the surface of the at least one wall is a matte surface or anembossed surface. Also, any of the preceding methods may comprisewherein the collapsible, flexible container(s) is pre-folded.

EXAMPLES

Tests were conducted to compare the vacuum integrity test ontraditional, flat or non-embossed film containers versus embossed filmcontainers. As shown in FIG. 3, non-embossed film with or without a 30μm defect was similar to the test result of embossed film with no defect(control). In contrast, the embossed film with a 30 μm defect wasreadily distinguishable from control within seconds of starting the test(start time point at approximately 98 seconds). By 102 seconds (e.g.,less than 4 seconds of test time), the defect in the embossed film 20 Lcontainer was conclusively identified.

FIG. 4 demonstrates that the vacuum integrity test is highly sensitive,finding a single 2 μm hole in a 20-liter bag manufactured using anembossed film, as described herein. The flow of air, in cubiccentimeters per minute, is significantly higher, and easily detectable,for the bag having a defect as small as 2 microns.

FIGS. 5 and 6 demonstrate that the increasing the size of the testcontainer to 200-liters and even 500-liters nonetheless positivelyidentifies defects as small as 2 microns in embossed film containers.Note that the initial increase in mass flow results from opening thevalve to the instrument.

FIG. 7 demonstrates that the vacuum integrity test positively identifiesdefects in embossed film containers post-use. As described above, it maybe desirable to test a flexible container after use to confirmintegrity. This test confirms that 50 μm defects were positivelyidentified in embossed film bags pre- and post-use as compared to anembossed film control without a defect pre- and post-use.

The present disclosure is not to be limited in scope by the specificembodiments described herein. Indeed, other various embodiments of andmodifications to the present disclosure, apparent to in addition thoseof to those described herein, ordinary skill in the art will from be theforegoing description and accompanying drawings. Therefore, such otherembodiments and modifications are intended to fall within the scope ofthe present disclosure. Furthermore, although the present disclosure hasbeen described herein in the context of a particular implementation in aparticular environment for a particular purpose, those of ordinary skillin the art will recognize that its usefulness is not limited thereto andthat the present disclosure may be beneficially implemented in anynumber of environments for any number of purposes. Accordingly, theclaims set forth below should be contemplated in view of the fullbreadth and spirit of the present disclosure as described herein.

What is claimed is:
 1. A method comprising: a) providing a collapsible,flexible bioprocessing container, the container comprising at least twoopposing flexible walls, wherein the surface of the walls internal tothe container comprise a plurality of channels on said walls; b)evacuating air from the container to a negative atmospheric pressure; c)monitoring mass flow so as to determine the integrity of the container.2. The method of claim 1, wherein the plurality of channels are embossedon said walls.
 3. The method of claim 1, wherein the surface of thewalls are matte surfaces.
 4. The method of claim 1, wherein thecollapsible, flexible bioprocessing container is pre-folded.
 5. Themethod of claim 1, wherein the collapsible, flexible bioprocessingcontainer further comprises fittings, tubing, connectors, and acombination thereof.
 6. The method of claim 1, wherein the negativeatmospheric pressure is between about −1 psi to −14 psi.
 7. A method ofdetecting a 2 micron or larger defect in a collapsible, flexiblebioprocessing container, the method comprising: a) providing acollapsible, flexible bioprocessing container, the container comprisingat least two opposing flexible walls, wherein the surface of the wallsinternal to the container comprise a plurality of channels on saidwalls; b) evacuating air from the container to a negative atmosphericpressure; c) monitoring mass flow, wherein an increase in mass flowindicates the presence of a defect.
 8. The method of claim 7, whereinthe plurality of channels are embossed on said walls.
 9. The method ofclaim 7, wherein the surface of the walls are matte surfaces.
 10. Themethod of claim 7, wherein the collapsible, flexible bioprocessingcontainer is pre-folded.
 11. The method of claim 7, wherein thecontainer further comprises fittings, tubing, connectors, and acombination thereof.
 12. The method of claim 7, wherein the negativeatmospheric pressure is between about −1 psi to −14 psi.
 13. Acollapsible, flexible bioprocessing container, the container comprisingat least two opposing flexible walls, wherein the surface of the wallsinternal to the container comprise a plurality of channels on saidwalls.
 14. The container of claim 13, wherein the plurality of channelsare embossed on said walls.
 15. The container of claim 13, wherein thesurface of the walls are matte surfaces.
 16. The container of claim 13,wherein the collapsible, flexible bioprocessing container is pre-folded.17. The container of claim 13, wherein the container further comprisesfittings, tubing, connectors, and a combination thereof.
 18. The methodof claim 8, wherein the surface of the walls are matte surfaces.
 19. Thecontainer of claim 14, wherein the surface of the walls are mattesurfaces.
 20. The container of claim 14, wherein the collapsible,flexible bioprocessing container is pre-folded.